Increased Ghrelin but Low Ghrelin-Reactive Immunoglobulins in a Rat Model of Methotrexate Chemotherapy-Induced Anorexia - PubMed (original) (raw)

doi: 10.3389/fnut.2016.00023. eCollection 2016.

Kuniko Takagi 1, Romain Legrand 1, Nicolas Lucas 1, Stephanie Beutheu 1, Christine Bôle-Feysot 1, Aurore Cravezic 2, Naouel Tennoune 1, Jean-Claude do Rego 2, Moïse Coëffier 3, Akio Inui 4, Pierre Déchelotte 3, Sergueï O Fetissov 1

Affiliations

Increased Ghrelin but Low Ghrelin-Reactive Immunoglobulins in a Rat Model of Methotrexate Chemotherapy-Induced Anorexia

Marie François et al. Front Nutr. 2016.

Abstract

Background and aims: Cancer chemotherapy is commonly accompanied by mucositis, anorexia, weight loss, and anxiety independently from cancer-induced anorexia-cachexia, further aggravating clinical outcome. Ghrelin is a peptide hormone produced in gastric mucosa that reaches the brain to stimulate appetite. In plasma, ghrelin is protected from degradation by ghrelin-reactive immunoglobulins (Ig). To analyze possible involvement of ghrelin in the chemotherapy-induced anorexia and anxiety, gastric ghrelin expression, plasma levels of ghrelin, and ghrelin-reactive IgG were studied in rats treated with methotrexate (MTX).

Methods: Rats received MTX (2.5 mg/kg, subcutaneously) for three consecutive days and were killed 3 days later, at the peak of anorexia and weight loss. Control rats received phosphate-buffered saline. Preproghrelin mRNA expression in the stomach was analyzed by in situ hybridization. Plasma levels of ghrelin and ghrelin-reactive IgG were measured by immunoenzymatic assays and IgG affinity kinetics by surface plasmon resonance. Anxiety- and depression-like behaviors in MTX-treated anorectic and in control rats were evaluated in the elevated plus-maze and the forced-swim test, respectively.

Results: In MTX-treated anorectic rats, the number of preproghrelin mRNA-producing cells was found increased (by 51.3%, p < 0.001) as well were plasma concentrations of both ghrelin and des-acyl-ghrelin (by 70.4%, p < 0.05 and 98.3%, p < 0.01, respectively). In contrast, plasma levels of total IgG reactive with ghrelin and des-acyl-ghrelin were drastically decreased (by 87.2 and 88.4%, respectively, both p < 0.001), and affinity kinetics of these IgG were characterized by increased small and big Kd, respectively. MTX-treated rats displayed increased anxiety- but not depression-like behavior.

Conclusion: MTX-induced anorexia, weight loss, and anxiety are accompanied by increased ghrelin production and by a decrease of ghrelin-reactive IgG levels and affinity binding properties. Such changes of ghrelin-reactive IgG may underlie their decreased ghrelin-transporting capacities compromising ghrelin orexigenic and anxiolytic effects and contributing to chemotherapy-induced loss of appetite.

Keywords: anorexia; autoantibodies; chemotherapy; ghrelin; intestinal inflammation.

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Figures

Figure 1

Figure 1

Body weight (A), daily food intake (B), and water intake (C) of MTX-treated and PBS-treated control rats (both, n = 10). Ratios of daily water to food intakes (D). Two-way RM ANOVA with Bonferroni post-tests ***p < 0.001, effects of time p < 0.0001 (mean ± SEM).

Figure 2

Figure 2

Representative microphotographs of preproghrelin mRNA-expressing cells in the stomach of PBS-treated control rats (n = 10) (A) and MTX-treated rats (n = 7) (B) revealed by in situ hybridization with DIG-labeled preproghrelin anti-sense riboprobes. (C) Quantification of the number of preproghrelin mRNA-positive cells in the gastric mucosal and submucosal layers of control and MTX-treated rats. Student’s _t_-test, ***p < 0.001, KS test _p_ > 0.1 both (mean ± SEM). Scale bar, 100 μm.

Figure 3

Figure 3

Plasma concentrations of ghrelin (A), des-acyl ghrelin (B), and their ratios (C) in PBS-treated controls (n = 8) and MTX-treated (n = 10). *p < 0.05, Mann–Whitney test, KS test _p_ = 0.03 and _p_ > 0.1 and **p < 0.01 Student’s _t_-test, KS test _p_ > 0.1 both (mean ± SEM).

Figure 4

Figure 4

Plasma concentrations of free (A,D) and total (B,E) IgG reactive with ghrelin (A,B) and des-acyl ghrelin (D,E) in MTX-treated (n = 10) and PBS-treated control rats (n = 10). Mann–Whitney or Student’s _t_-test (A,E) according to the KS normality tests, *p < 0.05, **_p_ < 0.01, ***_p_ < 0.001. KS tests, **(A)** _p_ > 0.1 and p = 0.08, (B) p > 0.1 and p = 0.03, (C) p > 0.1 and p = 0.003, (D) p > 0.1 and p = 0.005, (E) p > 0.1 and p = 0.1, (F) p > 0.1 and p = 0.006 (mean ± SEM).

Figure 5

Figure 5

SPR affinity kinetics analysis of ghrelin- (A–C) and des-acyl ghrelin (D–F)-reactive IgG in MTX-treated (n = 9) and PBS-treated control rats (n = 7). The affinity kinetic parameters are: the association rates (small ka) (A,D), the dissociation rates (small kd) (B,E) and the dissociation equilibrium constants (KD) (C,F). Student’s _t_-test (B) or Mann–Whitney test according to the KS normality tests, *p < 0.05. KS tests, **(B)**, _p_ > 0.1 both and (F), p = 0.02 and p > 0.1 (mean ± SEM).

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